Calcium pyrophosphate dihydrate (CPPD) crystal deposition disease is a common form of arthritis, particularly in the elderly. Prevalence approaches 50 percent in those over 80. CPPD crystals cause acute attacks of gout-like arthritis, but more importantly are associated with debilitating degenerative arthritis. In vitro and in vivo data strongly suggest that these crystals cause or amplify cartilage degeneration. This proposal focuses on the source of inorganic pyrophosphate (PPi), the anion constituent of CPPD crystals. Disordered PPi metabolism is clearly implicated in CPPD crystal deposition. Specific emphasis will be placed on mechanisms underlying PPi generation in extracellular cartilage matrix where crystals form. The effector arm of extracellular PPi (ePPi) generation involves the ectoenzyme nucleoside triphosphate pyrophosphohydrolase (NTPPPH), which generates ePPi from extracellular nucleoside triphosphates such as ATP. NTPPPH activity is highly expressed in articular cartilage. The availability of extracellular ATP substrate for NTPPPH is rate-limiting for generation of ePPi and the most likely cellular source of that ATP is the chondrocyte. These studies are designed to determine the effects of factors that modulate ePPi formation upon the release of ATP from chondrocytes. Specific emphasis is placed on growth factors (transforming growth factor-beta and insulin-like growth factor-I), on chondrocyte donor age, on transduction pathways (protein kinase C and cAMP-related), and on purine receptors (P1and P2). All of the aforementioned influence ePPi elaboration by chondrocytes. The second goal of this proposal is to determine the mechanisms of ATP egress from chondrocytes. Specific focus will be on ATP binding cassette proteins (analogues of human ABC1 and p-glycoprotein expressed in chondrocytes), a gap junction protein (connexin 43), and ANK protein as possible transporters. The intent of these studies is to develop insights into the pathogenesis of CPPD deposition disease so that therapeutic options may be generated.